THE PRIMER PROTEIN OF BACTERIOPHAGE M2 INHIBITS ELONGATION ACTIVITY OF ITS DNA POLYMERASE

Abstract

The primer protein (PP) and DNA polymerase (Pol) of bacteriophage M2, both of which are essential for the protein-primed replication of the genome, were purified from Escherichia coil cells that harbored the recombinant plasmid pIKM23, on which the genes for PP and Pol are under the control of the tac promoter. The purified PP and Pol formed a heterodimeric complex at a molar ratio of 1 to 1. Maximal activity of Pol for M2 DNA replication was attained when an equimolecular amount of PP, relative to that of Pol, was added to the replication system in vitro. When excess amount of PP was added, however, the replication activity was reduced. The activity of Pol for the priming reaction in M2 DNA replication was not reduced even by the presence of a five-fold molar excess of PP over Pol. By contrast, the activities for DNA chain elongation with templates of primed M2 DNA and with poly(dA): oligo (dT) were inhibited by 70% and by 80%, respectively, on the addition of PP in an equimolecular amount to that of Pol. Therefore, the reduction in M2 DNA replication appears to be caused by an inhibitory effect of PP on Pol in the elongation process.